Positional cloning (also known as long-range cloning) refers to the process of identifying a gene by first determining its chromosomal location, and then isolating and screaming candidate genes from that region to isolate the specific gene. In the positional cloning projects described in this program, the chromosomal locations of the human and mouse loci of interest have been determined in the individual projects using genetic mapping techniques. Once the map position is sufficiently narrow, the strategy for isolation of the underlying gene is largely similar for the different genes, and is outlined in Fig.1. First, the region defined by genetic mapping is isolated in the form of large insert DNA clones. This is accomplished by screening libraries that carry large genomic DNA inserts, typically in the form of YAC (yeast artificial chromosome), BAC (bacterial artificial chromosome), and PAC (bacteriophage PI artificial chromosome) clones. Next, physical mapping of the clones is performed to determine clone insert size, overlap between inserts, and interclone distance. The goal is to cover the critical gene region with contiguous overlapping clones to produce a DNA 'contig' that contains the gene of the critical gene region with contiguous overlapping clones to produce a DNA 'contig' that contains the gene of interest. Various methods may then be employed to screen individual clones in the contig for the presence of the gene. The strategy used at this stage is dependent on properties of the specific mutation; one of the most powerful approaches, which will be used in multiple projects, is complementation of the mutant phenotype. Once a single genomic clone has been identified, efforts focus on identification of transcribed gene sequences using techniques such as direct cDNA selection, exon trapping and random sequencing. Finally, candidate genes identified from such as direct cDNA selection, exon trapping and random sequencing. Finally, candidate genes identified from the genomic region are evaluated by sequencing the corresponding gene to identify the mutation at the nucleotide.